Methods and compositions for treating and preventing mucositis

ABSTRACT

A method of reducing or inhibiting mucositis in a patient, which includes administering an inflammatory cytokine inhibitor or a mast cell inhibitor, or a combination thereof, is disclosed.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of Ser. No. 09/800,855 filed Mar. 7,2001, which is a continuation of Ser. No. 09,265,299 filed Mar. 9, 1999,now U.S. Pat. No. 6,458,777 B1, which is a continuation-in-part of U.S.Ser. No. 09/065,012, filed Apr. 23, 1998, now abandoned, which claimspriority from United States provisional patent application No.60/077,977, filed Mar. 13, 1998.

BACKGROUND OF THE INVENTION

This invention relates to methods and compositions for treating andpreventing mucositis.

Mucositis is the destruction of the oral mucosal epithelium, whichresults in erythema, ulcerations and severe pain in the oral cavity.Mucositis often arises as a complication of antineoplastic therapy, suchas cancer chemotherapy and/or radiation therapy. The painful ulcerativelesions of mucositis can cause patients to restrict their oral intake;as a result, they lose weight and suffer from fever associated withdehydration. Severe mucositis can necessitate the de-escalation of aplanned chemo/radio-therapeutic dosing regimen to prevent further damageto the oral mucosa.

An even more serious consequence of mucositis is that the lesions canact as sites of secondary infections and as portals of entry forendogenous oral microorganisms. Mucositis is therefore a significantrisk factor for life-threatening systemic infection (septicemia); therisk of systemic infection is exacerbated by concomitant neutropenia,which is another complication associated with chemotherapy. Patientswith mucositis and neutropenia have a relative risk of septicemia thatis at least four times greater than that of individuals withoutmucositis.

The overall frequency of mucositis varies; it is influenced by thepatient's diagnosis, age, and level of oral health, as well as the type,dose, and frequency of drug or radiation administration. Approximately40% of all patients who receive cancer chemotherapy suffer some degreeof mucositis, and virtually 100% of patients treated with radiationtherapy for head and neck tumors develop mucositis. The frequency ofsevere mucositis in patients undergoing high risk protocols is over 60%.About 50% of individuals develop lesions severe enough to requiremodification of their cancer treatment and/or parenteral analgesia.

The development of effective methods for treating and preventingmucositis has been hampered by a lack of understanding of thepathophysiology of this condition, and by the inconsistency in patientresponse to the medications currently in use.

SUMMARY OF THE INVENTION

The invention features methods for treating and preventing mucositis.The invention is based, in part, on the recognition that mucositis is acomplex biological process resulting from the cumulative and interactiveeffects of radiation and/or chemotherapy with epithelial connectivetissue and endothelium, pro-inflammatory cytokines, cellular elementswithin the mucosa and the local oral environment.

We hypothesize that mucositis represents a clinical outcome due to acomplex interaction of local tissue (connective tissue, endothelium,epithelium) toxicity, the level of myelosuppression and the oralenvironment. The local tissue components include an oral mucosa ofrapidly renewing stratified squamous epithelium overlying a loose andrichly vascular connective tissue base and appear to be responsive tochanges in patients' bone marrow status and, particularly, the degree ofgranulocytopenia. The oral microbial flora, saliva and functional traumaprovide an indigenous environment which impacts on the frequency,severity and course of chemotherapy-associated stomatotoxicity.

It is quite likely that the initial oral tissue response to chemotherapyand radiation occurs at the endothelial and connective tissue level. Webelieve that free radical formation leads to the disruption offibronectin with subsequent activation of transcription factors,stimulation of pro-inflammatory cytokine production and tissue damage. Arelationship between the presence of tumor necrosis factor-alpha (TNF-α)and IL-1 in serum correlates with the presence of non-hematologictoxicities. It is also likely that injury to endothelial cells occurssimultaneously. Concurrently, damage to the basal epithelial cellsprevents their replication. It is unclear whether many of these cellsundergo apoptosis or necrosis. An influx of inflammatory cellsexpressing pro-inflammatory cytokines occurs during the breakdown of themucosa and peaks just prior to the acme of mucositis. Bacterialcolonization of the damaged epithelium occurs and is accelerated by thepatient's myelosuppressed state. Typically the nadir follows a day or soafter peak mucositis. Bacterial cell wall products from both grampositive and gram negative organisms likely then penetrate the injuredmucosa and further stimulate the release of damaging cytokines. Finally,the mucosa recovers, a process which takes about three weeks in theabsence of secondary infection.

According to the invention, mucositis can be treated, or even prevented,by the administration of inflammatory cytokine inhibitors, MMPinhibitors, and/or mast cell inhibitors. The combination of theseinhibitors with an anti-inflammatory agent and/or an antimicrobial agentprovides an even more effective regime for preventing and treatingmucositis.

The invention features a method of reducing or inhibiting mucositis, ina patient suffering from mucositis or at risk for mucositis; the methodincludes administering to the patient a first therapeutic agent in anamount sufficient to inhibit mucositis, where the first therapeuticagent is an inflammatory cytokine inhibitor, a mast cell inhibitor, anMMP inhibitor, or a combination of these inhibitors. Preferred mast cellinhibitors include degranulation inhibitors, antihistamines, and serineprotease inhibitors. A preferred MMP inhibitor is a tetracycline such asminocycline, which used by itself in low doses is an effective mucositisagent that does not primarily act as an antibiotic. Other members of thetetracycline family can be used as well, e.g., chlortetracycline andoxytetracycline. An example of a mucositis that can be reduced orinhibited according to the invention is oral mucositis.

The invention also features a method of treating, inhibiting, orpreventing mucositis in the human patient by administering to thepatient first and second different therapeutic agents, the first agentbeing an NSAID (non-steroidal anti-inflammatory), an inflammatorycytokine inhibitor, or a mast cell inhibitor, and the second agent beingan inflammatory cytokine inhibitor, a mast cell inhibitor, an MMPinhibitor, an NSAID, or an NO inhibitor. Preferrably at least one of theagents is an NSAID, which is a COX-1 or COX-2 inhibitor; examples ofCOX-1 inhibitors are indomethacin and flurbriprofin. In other preferredembodiments, the first agent is an inflammatory cytokine inhibitorselected from an IL-6 inhibitor, a TNF-alpha inhibitor, an IL-1inhibitor, and an interferon-gamma inhibitor. A preferred combination isa TNF-alpha inhibitor combined with an MMP inhibitor such as atetracycline, eg, minocycline. Exemplary NO inhibitors areaminoguanidine and guanidine. Another TNF-alpha inhibitor that can beused is thalidomide. Mast cell inhibitors can be antihistamines, serineprotease inhibitors, or degranulation inhibitors.

In other preferred methods, a third therapeutic agent, in an amountsufficient to inhibit infection, is administered as well; the thirdtherapeutic agent includes an antimicrobial compound. Preferably, thefirst, second, and third therapeutic agents are administeredconcurrently.

In another preferred method, the first therapeutic agent, in an amountsufficient to inhibit mucositis, and the third therapeutic agent, in anamount sufficient to inhibit infection, are administered. Preferably,the first therapeutic agent and the third therapeutic agent areadministered concurrently.

The mucositis being treated can be induced by antineoplastic therapy;for example, it can be induced by chemotherapy or by radiation therapy.The patient treated with the methods and compositions of the inventioncan be a cancer patient preparing to undergo chemotherapy or radiationtherapy, or a cancer patient currently undergoing chemotherapy orradiation therapy.

The invention further features a pharmaceutical composition for treatingoral mucositis that includes (a) a first therapeutic agent including aninflammatory cytokine inhibitor, a mast cell inhibitor, an MMP inhibitoror a combination of these inhibitors; (b) a second therapeutic agentincluding an anti-inflammatory agent; and (c) a pharmaceuticallyacceptable carrier. The first and second therapeutic agents are presentin amounts sufficient to inhibit mucositis in a patient suffering frommucositis or at risk for mucositis. Preferably, the composition isformulated into a lozenge, a tablet, an oral rinse, an oral paste, or anoral gel. A preferred mast cell inhibitor is an antihistamine; preferredanti-inflammatory agents include non-steroidal anti-inflammatory drugsand cyclooxygenase-2 inhibitors. Preferred MMP inhibitors includetetracyclines such as minocycline, tetracycline HCl, or doxycycline.Preferred compositions can also include an anti-ulcer agent, in anamount sufficient to inhibit gastric mucosal injury, and anantimicrobial agent, in an amount sufficient to inhibit infection.

BRIEF DESCRIPTION OF THE DRAWING

FIG. 1 is a schematic representation illustrating the four phases ofmucositis development and resolution.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

The invention features methods and compositions for reducing andinhibiting mucositis that include administering inflammatory cytokineinhibitors and/or mast cell inhibitors.

The invention is based, in part, on the development of a new mechanisticscheme for the physiological basis of mucositis. According to thisscheme, the development and resolution of mucositis occurs in fourinterrelated phases: (i) an inflammatory/vascular response; (ii) adegenerative connective tissue and/or epithelial phase; (iii) anulcerative/bacteriological phase; and (iv) a healing phase. The fourphases are illustrated in FIG. 1.

During phase 1, the inflammatory or vascular phase, the administrationof chemotherapy effects the release of the cytokines interleukin-1(IL-1), interleukin-6 (IL-6), and tumour necrosis factor-alpha (TNF-α)from the epithelium. Alternatively, the administration of ionizingradiation causes the release of these cytokines from both the epitheliumand from the surrounding connective tissues.

IL-1 induces an inflammatory response that results in increasedsub-epithelial vascularity, with a consequent increase in the locallevels of cytotoxic agents. Both IL-1 and TNF-α cause local tissuedamage, and thereby initiate and accelerate mucositis.

During phase 2, the degenerative epithelial phase, radiation andchemotherapeutic drugs affect the endothelium, the connective tissuesand the dividing cells of the oral basal epithelium, resulting inreduced epithelial renewal, atrophy, and ulceration. The ulceration ofthe surrounding tissue is exacerbated by functional trauma and by aflood of locally produced cytokines.

Phase 3, the ulcerative/bacterial phase, is the most symptomatic andperhaps the most complex. This phase generally occurs at the time of thepatient's maximum neutropenia. Phase 3 is characterized by the releaseof agents that stimulate cytokine production from bacteria on thelesions. Localized areas of full-thickness erosion develop, and afibrous pseudomembrane sometimes grows over these areas. Secondarybacterial colonization of the lesions occurs, including colonizationwith both gram positive and negative organisms; this stimulates cytokinerelease from the surrounding connective tissue, which further amplifieslocal tissue destruction.

During phase 4, the healing phase, epithelial proliferation anddifferentiation is renewed, the peripheral white blood cell count isnormalized, and the local microbial flora is re-established.

These four phases are interdependent; they are the consequence of aseries of actions mediated by cytokines, the direct effect of theantineoplastic agents on the epithelium, connective tissue andendothelium, the oral bacterial flora, and the status of the patient'sbone marrow.

The invention is also based, in part, on the discovery thatproliferation of mast cells plays a key role in the development ofmucositis. Mast cells are granule-containing secretory cells which arepresent in mucosal and connective tissues, and which can migrate withinthese tissues. The distribution of mast cells in tissues generallyrelates to the potential of mast cell-derived mediators to influencecells in the immediate environment. In the oral cavity, mast cells arepreferentially distributed within the microvascular bed of the mucosa.

The granules of mast cells contain mediators that promote inflammation.Following degranulation, which can be triggered by a variety of stimuli,such as IgE, neuropeptides, trauma, and drugs, the mast cell mediatorsare deposited in large quantities in the extracellular environment.These mediators include histamine; the serine proteases chymase andtryptase; and cytokines, including TNF-α. The mediators promoteinflammation by exerting their effects on endothelial cells and othercell types. For example, the mediators may influence adhesion moleculesand the behavior of the tissue, leading to ulceration.

Two of the most important of these mediators are histamine and TNF-α. Inthe normal oral mucosa, these mediators are present only in the granulesof mast cells, and are absent in other cells.

Mast cell-released histamine increases vascular permeability byeffecting structural changes, such as endothelial contraction andintercellular gap formation. These changes result in increased locallevels of chemotherapy-induced damage. In addition, histamine promotesleukocyte adhesion to endothelial cells via transient mobilization ofthe adhesion molecule, P-selectin, thereby causing inflammation.

Another important mediator released by mast cells is the cytokine TNF-α.TNF-α contributes to the inflammatory process by releasing histamine andby inducing endothelial expression of E-selectin, an adhesion moleculewhich is critically required for the rapid adhesion of neutrophils, Tcells, monocytes, and other leukocytes to endothelial cells.

According to the invention, agents that inhibit the function of the mastcells or the action of the mediators released by mast cells can be usedto treat and prevent mucositis. Mast cell inhibitors are chemical orbiological agents that suppress or inhibit the function of mast cells,or the mediators released by mast cells. For example, mast cellinhibitors can inhibit degranulation, thereby preventing the release ofmediators into the extracellular space. Examples of mast celldegranulation inhibitors include picetannol, benzamidines, tenidap,tiacrilast, disodium cromoglycate, lodoxamide ethyl, and lodoxamidetromethamine. Other agents that inhibit mediator release includestaurosporine and CGP 41251.

Examples of mast cell mediator inhibitors include agents that block therelease or secretion of histamine, such as FK-506 and quercetin;antihistamines such as diphenhydramine; and theophylline.

Other mast cell inhibitors include serine protease inhibitors, such asα-1-protease inhibitor; metalloprotease inhibitors; lisofylline; TNFR-FE(available from Immunex, Seattle, Wash.); benzamidine; amiloride; andbis-amidines such as pentamidine andbis(5-amidino-2-benzimidazolyl)methane.

According to the invention) inflammatory cytokine inhibitors can also beused to treat and prevent mucositis. Inflammatory cytokine inhibitorsare chemical or biological agents that suppress or inhibit inflammatorycytokines. Such inhibitors include pyridinyl imidazoles, bicyclicimidazoles, oxpentifylline, thalidomide and gabexate mesilate.

Anti-inflammatory agents can be used in combination with inflammatorycytokine and/or mast cell inhibitors to treat and prevent mucositisaccording to the invention. Examples of anti-inflammatory agents thatcan be used in the present invention include the non-steroidalanti-inflammatory drugs flurbiprofen, ibuprofen, sulindac sulfide, anddiclofenac. When NSAID's are administered according to the invention,anti-ulcer agents such as ebrotidine can be administered, e.g., to helpprotect against gastric mucosal damage

Other anti-inflammatory agents that can be used in the present inventioninclude misoprostil; methylxanthine derivatives, such as caffeine,lisofylline, or pentoxyfylline; benzydamine; naprosin; mediprin; andaspirin.

Another important class of anti-inflammatory agents includescyclooxygenase (COX) inhibitors, particularly COX-2 inhibitors. COX-2,an inducible enzyme stimulated by growth factors, lipopolysaccharide,and cytokines during inflammation or cell injury, is responsible for theelevated production of prostaglandins during inflammation. COX-2inhibitors are especially useful where the invention is used to treatmucositis in cancer patients undergoing chemotherapy or radiationtherapy, because of the gastrointestinal tolerability of theseinhibitors. COX-2 inhibitors that can be used in the invention includecelecoxib, nimesulide, meloxicam, piroxicam, flosulide, etodolac,nabumetone, and1-[(4-methylsulfonyl)phenyl]-3-trifluoromethyl-5-[(4-fluoro)phenyl]pyrazole.

Other useful anti-inflammatory agents include dualcyclooxygenase/lipoxygenase inhibitors, such as2-acetylthiophene-2-thiazolylhydrazone, and leukotriene formationinhibitors, such as piriprost.

MMP inhibitors include both the antibacterial tetracyclines such astetracycline HCl, minocycline and doxyocycline, as well asnon-antibacterial tetracyclines.

The presence of bacteria in the oral cavity leads to secondaryinfection, serves as a source for systemic infection, and stimulatescytokine release, thereby amplifying tissue damage. According to theinvention, the administration of anti-microbial agents in combinationwith the agents described above can result in an even more effectivemethod for treating and preventing mucositis. Examples of antimicrobialagents that can be used include agents with spectrum for gram positiveand gram negative organisms. Specific drugs include tetracycline,amoxicillin, gentamicin, and chlorhexidine.

Other agents that can be used to treat or prevent mucositis include thenuclear transcription factor kappa-B (NF-κB) activation inhibitorscapsaicin and resiniferatoxin.

Route and Timing of Administration

The route of administration is governed by the nature of the compound(s)used. For example, the compounds can be administered in tablet orlozenge form, as an oral rinse, as a paste or gel, or by parenteraladministration.

Since the compositions of the invention can help prevent mucositis,administration of the compositions should preferably precede the initialdose of antineoplastic therapy by at least 24 hours. Daily treatmentshould continue during the course of antineoplastic treatment.

Dosage

The therapeutic agents described above can be used in the dose rangescurrently used for these agents. For topical application, the amount ofdrug to be administered will produce local tissue dose ranges equivalentto, or higher than, those achieved by parenteral administration. Thefollowing are illustrative examples of dose ranges.

Mast Cell Function Inhibitors

The mast cell function inhibitor, picetannol, is preferably administeredto tissue or plasma levels of 0.1 μg/ml to 5 μg/ml; benzamidines arepreferably administered to tissue or plasma levels of 0.5 to 1.0 μM/l;tenidap is preferably administered to tissue or plasma levels of 1-200μM/l; and tiacrilast is administered in a 1% to 10% solution.

Mast Cell Mediator Inhibitors

With respect to mediator inhibitors, lisofylline is preferablyadministered at 1 mg/kg to 10 mg/kg body weight, and TNFR-Fe (Immunex,Seattle, Wash.) is administered in 25 mg doses, twice weekly.

Anti-Inflammatory Agents

The anti-inflammatory agent, ibuprofen, is preferably administered at 50mg to 800 mg per day, and flurbiprofen is preferably administered at 50mg to 300 mg per day. The COX-2 inhibitor etodolac is preferablyadministered at 500 to 2000 mg per day; nabumetone is preferablyadministered at 500 to 2000 mg per day; meloxicam is preferablyadministered at 7.5 to 25 mg per day; piroxicam is preferablyadministered at 10 to 30 mg per day; and1-[(4-methylsulfonyl)phenyl]-3-trifluoromethyl-5-[(4-fluoro)phenyl]pyrazoleis preferably administered at 1 to 10 mg/kg per day.

Anti-Microbial Agents

With respect to anti-microbial agents, tetracycline is preferablyadministered at 250 mg to 1000 mg per day, and chlorhexidine ispreferably administered in a 0.1 to 5% solution, twice daily.

Without further elaboration, it is believed that one skilled in the artcan, based on the description herein, utilize the present invention toits fullest extent. The following specific examples are, therefore, tobe construed as illustrative of the invention, and are not meant tolimit the remainder of the disclosure in any way. Publications mentionedherein are hereby incorporated by reference.

MMP Inhibitors

Tetracyclines used as MMP inhibitors will be administered topically indosages of 0.001 to 10 mg/mL, with a probable range of 0.01 to 1 mg/mL,and an optimal range of 0.05 to 0.5 mg/mL.

EXAMPLE 1 Prophylaxis and Treatment for Patients UndergoingMyeloablative Chemotherapy and Total Body Irradiation in Preparation forBone Marrow Transplantation

For treatment according to the methods described herein, patients aredosed with a topical application of mucositis medication as a troche orlozenge, beginning the evening before the first dose of chemotherapy.The lozenge contains therapeutic doses of an MMP inhibitor such asminocycline and a nonsteroidal anti-inflammatory agent such asflurbiprofen.

Beginning the day of chemotherapy and continuing for the subsequentfourteen days, patients receive medication every 3 to 4 hours whileawake. For patients unable to tolerate the lozenge because ofchemotherapy-induced nausea, a non-viscous liquid suspension isavailable for dosing approximate every 2 hours while awake. Patientsusing the suspension swish and gargle with the material to assureexposure of the drug to the oropharynx. The fourteen-day dosing periodprovides coverage through the first three phases of mucositisdevelopment.

EXAMPLE 2 Prophylaxis and Treatment for Patients Undergoing RadiationTherapy for Tumors of the Head and Neck

Patients being treated with radiation therapy for head and neck cancersreceive a total tumor dose of radiation of about 60 Gy, given in divideddoses over a 6-week to 8-week period. Early signs of mucositis are notedat doses of around 10 Gy, and frank breakdown of the mucosa is seen ataround 25 Gy.

Beginning with the second week of this type of radiation therapy,patients receive mucositis medication 2 hours prior to each daily doseof radiation, which is typically given 5 days per week. Subsequentmucositis medication is given 2 hours, 6 hours, and 12 hours followingdaily radiation. Since myelosuppression is not an issue for patientsbeing radiated for head and neck cancers, the mucositis preparationincludes mast cell inhibitors, cytokine inhibitors, andanti-inflammatory agents, but no anti-microbial agents. Patients do notreceive mucositis medication on days on which they are not radiated. Theprotocol is followed until radiation dosing is completed.

EXAMPLE 3 Prophylaxis and Treatment for Patients Undergoing ChemotherapyTreatment for Treatment of Colorectal Cancer

In treatments for colorectal cancer, patients typically receivemultiple, monthly cycles of chemotherapy. Because of the use of specificanti-cancer drugs for treatment of this form of tumor, this group ofpatients is at particular risk for developing mucositis. Patients inthis group begin dosing with mucositis medication two hours prior tochemotherapy administration. They continue taking mucositis medicationevery 4 hours, while awake, for at least the next 48 hours. The regimenis repeated for each dosing cycle. Generally, the formulation does notinclude an anti-microbial. However, for those patients demonstratingsignificant neutropenia, the formulation includes an anti-microbial, andtreatment time is extended to ten days.

Other Uses

The methods and compositions of the present invention can be used totreat and prevent conditions such as lichen planus and graft-vs-hostdisease, which have similar biological mechanisms to that of mucositis.

Other embodiments are within the scope of the following claims.

What is claimed is:
 1. A pharmaceutical composition for treating oralmucositis comprising (a) a first therapeutic agent comprising an NSAID,an inflammatory cytokine inhibitor, or a mast cell inhibitor; (b) asecond, different therapeutic agent comprising an inflammatory cytokineinhibitor, a mast cell inhibitor, an MMP inhibitor, an NSAID, or an NOinhibitor; and (c) a pharmaceutically acceptable carrier, wherein saidfirst and second therapeutic agents are present in amounts sufficient toinhibit mucositis in a patient suffering from mucositis or at risk formucositis due to chemotherapy or radiation.
 2. The pharmaceuticalcomposition of claim 1, wherein said composition is formulated into alozenge, a table, an oral rinse, an oral paste, or an oral gel.
 3. Thepharmaceutical composition of claim 1, wherein said mast cell inhibitoris an antihistamine.
 4. The pharmaceutical composition of claim 1,wherein said anti-inflammatory agent is an NSAID.
 5. The pharmaceuticalcomposition of claim 4, wherein said composition further comprises ananti-ulcer agent in an amount sufficient to inhibit gastric mucosalinjury.
 6. The pharmaceutical composition of claim 1, wherein saidinflammatory cytokine inhibitor is a cyclooxygenase-2 inhibitor.
 7. Thepharmaceutical composition of claim 1, wherein said composition furthercomprises an antimicrobial agent in an amount sufficient to inhibitinfection.